Title Page
Contents
Abstract 8
I. INTRODUCTION 10
II. MATERIALS AND METHODS 13
1. Methods of ex-vivo expansion and in-vitro evaluation of immune cells 13
A. Ex-vivo immune cell expansion 13
B. Phenotype and cytotoxicity analyses of ex-vivo-expanded immune cells 13
C. Cytotoxicity assay 14
2. Methods for clinical applications 14
A. Study design 14
B. Historical control group 15
C. Inclusion and exclusion criteria for participant recruitment 15
D. Endpoints 21
E. Ex-vivo expansion and activation of immune cells and method of administration 21
F. Assessment of safety and efficacy 21
G. Imaging analysis 21
H. Methods of pre-treatment transcriptomic analyses of tissues from adoptive immune cells treated patients 22
I. Public GBM database 23
3. Statistical analysis 23
III. RESULTS 25
1. Ex-vivo expansion and in-vitro evaluation of immune cells 25
A. Proportions of immune cells in expanded PBMCs 25
B. Protein levels of activating and inhibitory NK cells receptors 27
C. Cytotoxicity effect of immune cells towards K562 cells 30
D. Cytotoxicity of immune cells towards GBM cell lines 30
2. Results of clinical trial 33
A. General information about participants 33
B. Safety of adoptive immune cell therapy 41
C. Clinical efficacy 47
D. Evaluation of tumor response by MRI 53
3. Using the NanoString assay to assess the transcriptomic landscape to identify the mechanism of immune cells 58
A. NanoString results revealed high immune reaction signatures within good responders 58
B. Expression of significantly upregulated genes in good responders or protective effects for PFS resembled a mesenchymal sub-type feature in GBM from TCGA dataset 65
C. Immunohistochemical staining of pre- and during Tx in a good responder 67
IV. DISCUSSION 69
V. CONCLUSION 74
REFERENCES 75
ABBREVIATIONS 80
ABSTRACT IN KOREAN 81
Table 1. Inclusion and exclusion criteria 18
Table 2. Patient treatments, status, and survival information of immune cell therapy group. 19
Table 3. Detailed relevant demographic of participants. 20
Table 4. Characterization of ex-vivo-expanded autologous immune cells from patients. 35
Table 5. Adoptive immune-cell therapy-related adverse events 42
Table 6. Total adverse events during clinical trial of immune cell therapy 43
Table 7. Clinical information of good responders and poor responders 49
Table 8. Patients treatments, status and survival information with historical control 51
Table 9. Genes associated with progression free survival identified by Cox regression or... 60
Table 10. Annotation-based unsupervised evaluation of label of clustering of genes significantly associated with PFS and label with good and poor responders 63
Fig 1. Characterization of ex-vivo-expanded immune cells. 26
Fig 2. Characterization of NK ligands in ex-vivo-expanded immune cells from seven... 28
Fig 3. Characterization of intracellular cytotoxicity-related proteins in... 29
Fig 4. Cytotoxicity assay of ex-vivo-expanded immune cells against K562 cell line. 31
Fig 5. Cytotoxicity assay of ex-vivo-expanded immune cells against GBM cell lines. 32
Fig 6. Participant enrollment flow diagram. 34
Fig 7. Therapeutic response assessment of the first recurrent GBM patient (A1) with the best prognosis (overall survival: 76 months)... 48
Fig 8. Kaplan-Meier plots. 50
Fig 9. MRI of A14 patient who did not receive treatment more than four times due... 54
Fig 10. Therapeutic response assessment of recurrent GBM A11 patient who showed... 54
Fig 11. T1-enhanced MRI after GTR or NTR of patients with poor prognosis. 55
Fig 12. T1-enhanced MRI after STR (A) or biopsy (B) patients with poor... 56
Fig 13. MRI of A6 patient. 56
Fig 14. Systemic demographic of radiographic tumor response assessment in participants 57
Fig 15. Proportions of regression risk (hazard ratio〉1) and protective(hazard ratio... 59
Fig 16. Volcano plot of DEG analysis of good and poor responders. 59
Fig 17. Cluster maps of four annotation terms (A-D) that fully discriminated... 64
Fig 18. Transcriptomic landscape in GBM patients from the TCGA dataset with... 66
Fig 19. Comparison of immune infiltration between pre-Tx (GTR) and during Tx... 68